Non-small cell lung cancer (NSCLC) is the leading cause of cancer mortality worldwide. In recent years dramatic progress has been made in tailoring therapies for subgroups of patients harboring specific genomic alterations, such as EGFR tyrosine kinase inhibitors for the 10-15% of patients bearing EGFR mutations, and through the use drugs blocking the PD-1/PD-L1 immune checkpoint pathway. Unfortunately, only a minority of patients benefit from these approaches. LKB1 (STK11) is the second most commonly altered tumor suppressor in NSCLC, and is lost in 20-30% of lung adenocarcinoma, resulting in 30,000-40,000 deaths annually. There is a major unmet need for therapeutic strategies tailored for LKB1-deficient (LD) NSCLC. Project investigators have demonstrated that LKB1 loss is associated with increased metastatic potential, chemotherapy resistance, and, more recently, with an immunosuppressed phenotype as well as resistance to checkpoint inhibitors. Given our initial findings, we hypothesize that a) LKB1 loss directly drives a distinctve immunosuppressed phenotype, and that potential underlying mechanisms include reduced antigen presentation and/or altered cytokine production; and b) therapeutic regimens can be developed to enhance the antitumor immune response and overcome resistance to checkpoint inhibition. We will test these hypotheses in the following aims. In Aim 1, we will characterize the immunosuppressed phenotype of LD-NSCLC, by a) investigating the mechanisms underlying the LD-associated intratumor immunosuppression in preclinical models, including reduced antigen presentation and altered production of immunosuppressive cytokines such as IL-6 and VEGF; and b) comparing the immune phenotype in LD and LKB1-intact (LI) tumors from NSCLC patients. Next, in Aim 2, we will use insights gained from Aim 1 to develop more effective immunotherapy approaches, by testing a) direct and indirect cytokine suppression, b) combinations of cytokine suppression with anti-PD1, to determine whether we can overcome the LD-associated resistance to checkpoint inhibition; and c) combinations with radiotherapy (RT) and other approaches enhancing antigen presentation. Finally, in Aim 3, we will translate this work into the clinic using a recently activated randomized clinical trial testing the anti-PD- antibody pembrolizumab, alone or combined with RT in 104 NSCLC patients. This will enable us to test our preclinical observations regarding the relative resistance of LD-NSCLC to PD-1 inhibition, and determine whether RT can enhance anti-tumor immunity and overcome PD-1 inhibitor resistance in LD NSCLC patients. Clinical significance: LD-NSCLC causes more deaths than pancreatic cancer, and there are critical unmet needs for new treatment approaches and insights into its distinct biology. We have assembled a multidisciplinary team of leading investigators to tackle these needs, with deep expertise in lung cancer genomics, immunotherapy, pathology, mouse models and radiotherapy that is poised to rapidly translate discoveries directly into clinical advances for NSCLC patients.